This application is being submitted in response to PAR-14-279 titled Discovery of in vivo Chemical Probes. The nuclear receptor (NR) superfamily of ligand regulated transcription factors has proven to be a rich source of targets for the development of therapeutics for a wide range of human diseases. The NR1F subfamily known as the retinoic acid receptor-related orphan receptors or RORs; ROR?(NR1F1), ROR?(NR1F2) and ROR?(NR1F3), regulate a wide range of physiological processes, including glucose and lipid metabolism, and immune functions. ROR? and ROR? are widely expressed in many tissues including liver, thymus, muscle, and skin while ROR? has a more restricted expression pattern and is found in regions of the central nervous system (CNS). While the identity of their physiological ligands remains controversial, we were the first to report sterols and oxygenated sterols as high affinity endogenous ligands and others have confirmed our findings and provided key evidence that they are physiological ROR ligands. The T cell specific ROR? isoform (ROR?t or ROR?2) has been shown to be the key lineage-defining transcription factor to initiate the differentiation program of TH17 cells making ROR?t the master regulator for TH17 and Tc17 differentiation, cells that have demonstrated anti-tumor efficacy, and ROR?t controls gene programs that enhance immunity (including increased IL17 production) and decrease immune suppression. Relevant to this proposal, we have exciting new data showing that activation of T cells using a synthetic agonist of ROR?t (SR1078) drives proliferation of TH17 cells and decreased expression of the immune checkpoint protein PD-1 (programmed cell death protein 1), a mechanism that will enhance anti-tumor immunity while blunting tumor associated adaptive immune resistance. Interestingly, putative endogenous sterols drive proliferation of TH17 cells but do not repress PD-1 expression. This suggests that synthetic agonists of ROR?t will activate TC17/TH17 cells (decreasing the population of Tregs), repress PD-1, and produce IL17 in situ (IL17 is associated with good prognosis in cancer). Enhanced immunity and blockage of immune checkpoints has transformed cancer treatment, thus such a molecule would provide a unique combination therapy with approved anti-PD-1 molecules for treatment of cancer. Importantly, our team was the first to identify synthetic modulators of the RORs. We developed the pan ROR?/ROR?t inverse agonist SR1001 and the potent ROR?t isoform selective inverse agonist SR2211, a compound that suppresses TH17 cell differentiation and blocks IL17 production. Critical to this proposal, we were the first to publish a synthetic pan ROR?/?t agonist, SR1078, capable of increasing of ROR target genes in HepG2 cells and in livers from mice treated with SR1078. Our labs have collaborated extensively to develop pan- ROR modulators, ROR?-selective agonists and inverse agonists, ROR?t-selective inverse agonists, and ROR?-selective inverse agonists. Thus we are well positioned to identify, characterize and develop potent and selective ROR?t agonists. Our goal is to optimize current pan scaffolds into ROR?t-selective agonists with good potency, physicochemical properties and pharmacokinetics to provide ROR?t agonists as probes to interrogate activation of ROR?t in vivo and determine its role in protective immunity.